Damage caused to tissues during ischemia/reperfusion can be extensive. Tissues deprived of oxygen suffer both reversible and irreversible damage. Injured tissues can also display disorders in automaticity. For example, myocardial tissues damaged during ischemia/reperfusion can display irreversible damage or myocardial infarction. Reversible damage, or stunning, is apparent with reduced pump efficiency leading to decreased cardiac output and symptomatology of suboptimal organ perfusion. Reperfusion of ischemic myocardial tissue may also cause electrophysiologic changes causing disorders in automaticity, including lethal arrythmias.
The exact mechanisms by which tissues are damaged during ischemia/reperfusion are unknown. It is hypothesized, however, that a complex series of events occur where tissues are damaged during ischemia as well as during subsequent reperfusion. During ischemia, tissues are deprived of oxygen-giving blood leading to anaerobic metabolism and consequently intracellular acidosis. Lack of circulation can cause infarcts or areas of necrotic, dead tissue. Ischemic tissues produce less of the enzymes needed to scavenge free radicals. Upon reperfusion and re-exposure to oxygen, tissues are damaged when free radicals including hydroxyl radicals are produced. Oxidative damage also disrupts the calcium balance in surrounding tissues causing stunning. Damage due to the oxidative burst is further compounded when injured cells release factors which draw inflammatory neutrophils to the ischemic site. The inflammatory cells produce enzymes which produce more toxic free-radicals and infiltrate the interstital spaces where they kill myocytes.
Methods to protect against the damage due to ischemia/reperfusion injury focus on reducing the initial oxidative burst and ensuing calcium overload preventing subsequent inflammation-associated damage. For example, agents which either decrease the production of oxygen-derived free radicals (including allopurinol and deferroxamine) or increase the catabolism of these materials such as superoxide dismutase, catalase, glutathione, and copper complexes, appear to limit infarct size and also may enhance recovery of left ventricular function from cardiac stunning. Agents which block sarcolemmal sodium/hydrogen exchange such as amiloride prevent the obligatory influx of calcium into the cell attendant with sodium extrusion and consequently reduce calcium overload.
Tissues can also be protected from ischemia/reperfusion injury by ischemic preconditioning. Ischemic preconditioning is triggered by brief antecedent ischemia followed by reperfusion which results in the rapid development of ischemic tolerance. This acute preconditioned state of ischemic tolerance lasts 30 min to 2 h and in myocardial tissue is characterized by reduced infarct size and a reduced incidence of ventricular arrythmias but not reduced levels of stunning (Elliot, 1998). Following dissipation of the acute preconditioned state, even in the absence of additional periods of preconditioning ischemia, a delayed preconditioned state of ischemic tolerance appears 12-24 h later and lasts up to 72 h. During the delayed phase of preconditioning protection against myocardial infarction, stunning and arrhythmia have been reported in various species.
Features of preconditioned myocardium in the face of ischemia/reperfusion include preservation of adenosine triphosphate (ATP) in some models, attenuation of intracellular acidosis and the reduction of intramyocyte calcium loading. Certain chemical agents known to be released by myocardium during ischemia have been shown to induce acute and delayed ischemic tolerance and provide cardiac protection. For example, adenosine, bradykinin and opiate receptor agonists which induce acute preconditioning appear to protect from ischemic injury via ATP dependent potassium (KAT) channel signaling pathways. The agent, bimakalim, known to open the KATP channel has also been shown to limit infarct size (Mizumura et al., 1995). Monophosphoryl lipid A (MLA) prevents irreversible as well as reversible damage to ischemic tissues (Elliot U.S. Pat. No. 5,286,718). Monophosphoryl lipid A is a detoxified derivative of lipid A, the active substructural element of lipopolysaccaride (LPS). LPS or endotoxin is a potent immunomodulator produced by most strains of Gram-negative bacteria. Pretreatment with LPS prior to ischemia has been shown to increase myocardial catalase activity increasing myocardial function (Brown et al., Bensard et al.). Endotoxin also protects against lung injury during hypoxia (Berg et al.). The cardioprotective effect of high doses of endotoxin appears to be associated with the ability of this xe2x80x9ctoxinxe2x80x9d to induce upon pretreatment myocardial oxidative stress, thereby protecting from a second oxidative stress associated with ischemia (Maulik et al.). LPS however is quite toxic. MLA has been structurally modified to negate the toxicity of LPS. It is hypothesized that MLA protects against injury due to ischemia/reperfusion injury by inducing the production of nitric oxide synthase which leads to an enhanced open-state probability of the cardioprotective ATP-dependent potassium channel (KATP). The nitric oxide burst caused by MLA may also lead to a decrease in the number of inflammatory neutrophils entering the post-ischemic area protecting the patient from further injury. In contrast to endotoxin, MLA does not appear to induce myocardial oxidative stress at cardioprotective doses.
Current treatments for ischemia/reperfusion injury are not however without drawbacks. Many of the agents known to be active, do not have broad clinical applicability, have limited effectiveness, and/or have dose limiting toxicities and consequently have been restricted in their application to ameliorate ischemia/reperfusion injury in the heart. Endotoxin is highly toxic to the system at cardioprotective doses. MLA, while non-toxic, is manufactured by the fermentation of S. minnesota and, as is the case with many biological products, exists as a composite or mixture of a number of molecular congeners varying in fatty acid substitution patterns with varying fatty acid chain lengths.
Although in comparison with endotoxin, MLA is non-toxic at cardioprotective doses, MLA can cause mild, transient, although not dose-limiting, fever in the target dose range. It should therefore be apparent from the above that a need remains for new compositions which are safe, effective and which have a broad clinical applicability in preventing or ameliorating the harmful effects of ischemia/reperfusion. Compositions which are non-toxic, non-pyrogenic, produced by chemical synthesis and of a single defined molecular structure would prove advantageous for this application.
The subject invention concerns a novel phosphoglycolipid useful in ameliorating the damage due to ischemia/reperfusion injury. Specifically, the claimed phosphoglycolipid is 2-Deoxy-6-O-[2-deoxy-4-O-phosphono-3-O-[(R)-3-tetradecanoyloxytetradecanoyl]-2-[(R)3-octadecanoyloxytetradecanoylamino]-xcex2-D-glucopyranosyl]-2-[(R)-3-hexadecanoyloxytetradecanoylamino]-D-glucopyranose and has the following structure: 
The phosphoglycolipid compound of the subject invention can be chemically synthesized using protocols described in the subject application.
Methods for using this novel phosphoglycolipid to ameliorate the damage due to ischemia/reperfusion injury are also described. The compound of the subject invention is particularly advantageous for this application because it does not induce fever or proinflammatory cytokines at dose levels anywhere near the cardioprotective dose range which may substantially increase the maximum tolerated dose and therefore the likelihood of achieving a therapeutic dose level in humans especially when administered immediately prior to surgery.